New aspects in biomass wastes valorization: spruce bark as plants growth regulators Presenting Author: Talmaciu, A. I.1, Co-Authors: Tanase, C.2, Bâra, C.I.3, Volf, I., Popa, V.I1 1“Gheorghe Asachi” Technical University of Iasi, Faculty of Chemical Engineering and Environmental Protection, D. Mangeron 73, 700050 Iasi, Romania 2 ‘’University of Medicine and Pharmacy” of Tirgu Mureș, Gheorghe Marinescu 38, 540139, Târgu Mureș, Mureș, Romania 3 “Alexandru Ioan Cuza” University, Faculty of Biology, 20 A, Bvd. Carol I, Iași 700506, Romania

1. Introduction – short background Table 1: Wastes generation UE* Waste (tones) Total waste Chemical and medical Animal and vegetal Mixed ordinary wastes European Union 2.515.220.000 57.880.000 110.060.000 282.010.000 Turkey 1.013.225.938 1.335.719 807.984 30.960.121 Germany 368.022.172 37.283.685 14.086.695 44.347.058 France 344.731.922 33.735.163 11.181.262 38.097.587 Romania 266.975.602 5.500.147 18.211.593 6.325.401 United kingdom 241.100.639 40.530.352 10.497.316 41.613.383 Fig. 1: Biomass categories for bioenergy** *http://appsso.eurostat.ec.europa.eu ** Gavrilesu M., 2008, Biomass power for energy and sustainable development , EEMJ Vol.7, No.5, 617-640

1. Introduction – Aim and objectives Biomass waste (forestry) Valorization and minimization Conversion (Extraction) Bio-products Plant industry/science or agriculture Resources Target Via For Uses

Ocimum basilicum L. seeds 2. Methods and material Spruce bark Wood processing waste Humidity = 10.44 % dm = 0.6 mm Basil seeds SC. Agrosel SRL, Campia Turzii, Romania Sterilization, HCl 20 %, 5 min Picea abies bark Ocimum basilicum L. seeds

2. Methods and material Extraction methods Biological tests on basil seeds Hot water extraction 10 g spruce bark 125 mL distilled water Water bath Temperature 70 – 80°C Extraction time 45 min 4 repetitions Ultrasound assisted extraction Ultrasound water bath (35 kHz) Temperature 70°C Extraction time 15 min Seeds germination tests - Seed germination energy and capacity Biometrical and physiological tests Vegetative organ's elongation Biomass accumulation Photoassimilating pigments content Cytogenetic tests Mitotic index Chromosomal aberrations Histo-anatomical tests Internal structure

3. Results and discussions 3.1 Extract characterization Table 1: Detailed spruce bark extracts composition Spruce bark extract Organic matter content (g/L extract) TPC (mg GAE/g) TTC TFC (mg RE/g) TFnolsC Aqueous extract obtained by hot extraction* 0.42±0.03 5.20±0.45 1.64±0.51 0.22±0.01 0.08±0.01 Aqueous ultrasonic extract 0.72±0.11 6.57±0.70 1.30±0.02 1.18±0.19 0.84±0.06 *Tanase et al., 2013; RSD < 1.2 % (n = 3); TPC – total polyphenolic content; TTC - total tannins content; TFC – total flavonoids content; TFnolsC – total flavonols content. Table 2: Spruce bark phenolic solutions investigated as growth regulator on basil seeds Working solution Abbreviation Concentration (g/L) TPC (mg GAE/mL) aqueous extract obtained by hot extraction SB1 5 0.06±0.01 SB2 10 0.13±0.05 aqueous ultrasonic extract SB3 0.35±0.08 SB4 0.65±0.04

3. Results and discussions 3.2 Seeds germination tests, vegetative organ’s elongation, biomass accumulation and photoasimilatory pigments content Fig. 1 Influence of spruce bark extracts on Ocimum basilicum L. seed germination energy and capacity Fig.2 Influence of spruce bark extracts on vegetative organs elongation of Ocimum basilicum L. Fig. 3 Influence of spruce bark extracts on biomass accumulation of Ocimum basilicum L.

3. Results and discussions 3.2 Seeds germination tests, vegetative organ’s elongation, biomass accumulation and photoasimilatory pigments content Table 3. The amount of photo-assimilatory pigments synthesized in Ocimum basilicum L. primary leaves Chl a µg/g Chl b Chl a+b Chl a/b Carotens C 123.68±2.23 29.96±0.51 153.64 4.12 62.22±0.73 SB1 133.98±2.47 38.72±0.63 172.69 3.46 80.42±0.86 SB2 96.63±1.45 26.08±0.22 122.71 3.70 88.46±0.81 SB3 127.94±2.65 34.84±0.55 162.78 3.67 83.41±0.79 SB4 108.76±2.01 25.11±0.21 133.87 4.33 61.83±0.53

3. Results and discussions 3.3 Cytogenetic test Fig.1 Mitotic index variation in root apex cells of Ocimum basilicum L. under the influence of tested solution Fig. 2 Frequent mitotic cell division phases in Ocimum basilicum L. root apex under the influence of tested solution

3. Results and discussions 3.3 Cytogenetic test – chromosomal aberrations a) Ana-telophase with late chromosomes b) Ana-telophase with expelled chromosomes c) Ana-telophase with multiple bridges

3. Results and discussions 3.4 Histo-anatomical tests Fig. 1 Internal structure of Ocimum basilicum L. root - control sample (absorbents bristle presence) Fig. 2 Internal structure of Ocimum basilicum L. root – spruce bark sample (SB2)

4. Conclusion

Future projects… …and acknowledgments Biological tests in culture vessels and land Antibacterial and antifungal activity tests on antibiotic-resistant strains Biological activity tests on plant cell culture and tissues …and acknowledgments Lect. Dr. Corneliu Tanase University of Medicine and Pharmacy Tirgu Mures  Department of Fundamental Pharmaceutical Sciences Pharmaceutical Botany - Cell Biology Email corneliu.tanase@umftgm.ro Lect. Dr. Csilla Iuliana Bara “ Alexandru Ioan Cuza” University of Iasi Faculty of Biology Department of Biochemistry and Molecular Biology, Iaşi Email csiulia@yahoo.com Assc. Prof. Dr. ing. Irina Volf “Gheorghe Asachi” Technical University of Iași, Faculty of Chemical Engineering and Environmental Protection, Department of Environmental Protection and Management Email iwolf@tuiasi.ro Prof. Dr. ing. Valentin i. Popa “Gheorghe Asachi” Technical University of Iași, Faculty of Chemical Engineering and Environmental Protection, Department of Natural and Synthetic Polymers Email vipopa@tuiasi.ro